The overarching goal of this project is to develop molecular tracers for imaging neuronal voltage changes deep in the brain. Photoacoustic imaging holds great promise to analyze the structure and function of neural circuitry deep in the brain, yet no compatible tools have been developed to probe neuronal activity. Therefore, we bring together unique resources in molecular tracer engineering and tissue-penetrating photoacoustic imaging technology to develop a toolbox of 'voltage tracers' for probing neural electrical activity. Neural circuitry is a dynamic network that incorporates neural activity across time and brain structures. Existing high-resolution light microscopy modalities, such as two-photon microscopy, combined with fluorescent reporters of neural activity allows simultaneous optical recordings from large populations of neurons in awake, behaving animals, especially rodents. However, these approaches permit functional visualization of single neurons only at depths of ?1mm, which is not even sufficient to image the entire thickness of the outermost layer of the brain, the cortex. To break light microscopy's 1mm barrier, photoacoustic tomography is a promising non-invasive technique that has enabled high-resolution imaging at mm to cm depths, with spatial resolution and coverage beyond the capability of fMRI and light microscopy, respectively. Photoacoustic imaging holds great promise for the visualization of physiological and pathogenic processes of deep in the brain. Yet, no biosensors of neural activity exist for photoacoustic imaging. This project will create the first neural activity probesfor photoacoustic imaging deep within the brain. Our goal is to develop and validate voltage tracers compatible with deep brain photoacoustic imaging of neuronal electrical signaling in vivo.